title: Turnover of beta-galactosidase in fibroblasts from patients with genetically different types of beta-galactosidase deficiency.
abstract: The turnover of lysosomal beta-galactosidase was studied in fibroblast cultures from patients with Gm1-gangliosidosis and combined beta-galactosidase and neuraminidase deficiency, which had 5-10% residual beta-galactosidase activity. beta-Galactosidase was specifically inactivated with the suicide substrate beta-D-galactopyranosylmethyl-p-nitro-phenyltriazene (beta-Gal-MNT) and from the subsequent restoration of enzyme activity in cell cultures turnover times were calculated. By using [3H]beta-Gal-MNT, the hydrolytic activity per molecule of beta-galactosidase was determined. 3H-labelled beta-D-galactopyranosylmethylamine, the precursor of [3H]beta-gal-MNT, was obtained by Raney-nickel-catalysed exchange with 3H2O. The rate of synthesis of beta-galactosidase in normal and all mutant cells tested was found to be 0.4-0.5 pmol/day per mg of cellular protein. The GM1-gangliosidosis cells tested contain the normal amount of 0.5 pmol of beta-galactosidase/mg of protein with a normal turnover time of about 10 days, but only 10% of beta-galactosidase activity per enzyme molecule. Cells with combined beta-galactosidase and neuraminidase deficiency contain only 0.3 pmol of beta-galactosidase/mg of protein with a decreased turnover time of 1 day and normal hydrolytic properties (200 nmol of 4-methylumbelliferyl galactoside/h pmol of beta-galactosidase).
mesh_terms: D001665:Binding Sites; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D005347:Fibroblasts / Q000201:enzymology*; D005696:Galactosidases / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D007787:Lactose Intolerance / Q000201:enzymology* / Q000235:genetics; D009439:Neuraminidase / Q000172:deficiency; D014226:Triazenes / Q000494:pharmacology; D001616:beta-Galactosidase / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D014226:Triazenes; C018547:beta-D-galactopyranosylmethyl-4-nitrophenyltriazene; D003513:Cycloheximide; D005696:Galactosidases; D009439:Neuraminidase; D001616:beta-Galactosidase